The ability of certain bacterial surface molecules to react selectively with constant regions of many classes and subclasses of IgG molecules from mammalian species has made these Fc-binding proteins of enormous value as immunochemical reagents. These binding proteins can be labeled to high specific activity or immobilized without loss of functional binding and can be used to detect and quantify antigens, fluid phase antibody, and antigen-antibody complexes. The utility of these reagents has been demonstrated by the large number of procedures developed using staphylococcal protein A and streptococcal protein G as tracers and immunoadsorbants for antibodies of the IgG isotype.
IgA is a class of antibody which is related to immunity against infections with bacteria and viruses at mucosal surfaces. It is present in virtually all mammalian secretions. Like other human antibodies, IgA is comprised of heavy and light chains, and is characterized by a constant fraction, Fc, and a variable fraction, Fab. The IgA antibody, like all antibodies, is produced by the lymphocytes of the immune system. To date, the availability of reagents that react selectively with antibodies of the IgA isotype, without interfering with the ability of the antibody molecule to bind to its cognate antigen has been extremely limited. For example, the IgA binding potential of the lectin jacalin is very limited because of its failure to react with both human IgA subclasses and by its non-specific interaction with non-IgA serum proteins (Bunn-Moreno, M. M., A. Campos-Neto [1981] J. Immunol. 127:427-429; Kondoh, H., K. Kobayashi, K. Hagiwara, T. Kajii [1986] J. Immunol. Methods 88:171-173).
Group B streptococci (GBS) are a class of microorganisms which has been extensively studied and classified. GBS are being increasingly recognized as important human pathogens. In addition to causing meningitis, bacteremia, endocarditis, bronchopneumonia, arthritis, peritonitis, wound infections, abscesses, and urinary tract infections in adults, as many as 80% of group B infections occur in neonates (Jelinkova, J. [1977] Current Topics in Microbiology and Immunology 76:127-165). Approximately 30% of pregnant women have been reported to be colonized by GBS. Despite this high carriage rate, neonatal infection occurs with an incidence of only 0.5%, resulting in over 12,000 deaths annually (Lim, D. V., Morales, W. J., Walsh, A. F., and Kazanis, D. [1986] J. Clin. Micro. 23:489-492). Predisposing factors to development of disease are premature birth, prolonged rupture of membranes, overt maternal infection, and deficiency of type specific antibody (Boyer, K. M. and Gotoff, S. P. [1986] New England J. Med. 314:1665-1669). It has now been discovered that certain of these streptococci, generally of the Ib or Ic serotype, will bind IgA.
Bacterial proteins with affinity for Ig classes other than IgG would be of considerable value as immunological tools. It is known that certain streptococcal strains bind IgA (Christensen and Oxelius [1975] Acta Path. Microbial. Scand. Sect. C, 83:184), and isolation of an IgA-binding protein from group B streptococci has even been reported (Russell-Jones et al. [1984] J. Exp. Med. 160:1467). See also U.S. Pat. No. 4,757,134. Western blot analysis of proteins extracted from these strains by treatment with detergent indicated that it may in fact be the p antigen component of the c protein marker complex which has the ability to bind to IgA (Russell-Jones, G. J. and Gotschlich, E. C. [1984] J. Exp. Med. 160:1476-1484). However, the extraction method used by this group-boiling of bacteria in 2% SDS--is not satisfactory for isolation of sufficient amounts of the protein, and the harshness of the procedure is likely to damage the protein. The protein is reported to have a molecular weight of 130 kDa.
In 1987 Cleat and Timmis reported that they had cloned a gene which codes for GBS beta antigen with ability to bind IgA (Cleat, P. H., K. N. Timmis [1987] Infect. Immun. 55:1151-1155). No nucleotide sequence has been reported for the DNA encoding the beta antigen. Recently, studies by Brady and Boyle have indicated that there are various forms of the beta antigen (Brady, L. J., M. D. P. Boyle [1989] Infect. Immun. 57:1573-1581). It was determined that there is a high molecular weight form bound to the surface of bacteria which binds to IgA. In addition, there are secreted proteins that exist in two forms, an IgA binding form and a non-IgA binding form.
In EPC patent application 87850160.0, an IgA-binding protein isolated from Streptococcus pyogenes strain AW 43 is described. EPC application 0 367 890 concerns a similar protein with similar binding characteristics but with a different N-terminal sequence. The proteins described in these European patent applications have been isolated from group A streptococci. It has been reported that the receptors obtained from group B streptococci are antigenically unrelated to the IgA receptor from group A streptococci (Lindahl, G. et al. [1990] Eur. J. Immunol. 20:2241-2247).
The subject invention pertains to the cloning and sequencing of a gene which codes for an approximately 45 kDa recombinant protein which binds with IgA.